Adenovirus vectors (AdVectors) designed to express an antigenic gene have shown great promise as vaccines. Although GenVec and other groups have constructed numerous AdVectors expressing a variety of antigens a substantial subset of antigen genes have been found to be inhibitory to AdVector construction and production. In this phase I SBIR proposal we will investigate strategies to circumvent the deleterious effect of inhibitory genes by blocking antigen expression within the AdVector production cell line. These results will guide the building, certification and banking of a commercial cell line during SBIR Phase II. A cell line broadly applicable for production of AdVectors containing inhibitory transgenes is presently not available. During SBIR phase I two different mechanisms of repressing expression, transcriptional and post-transcriptional, will be tested. Aim 1 will optimize a transcriptional repressor, incorporate it into a cell line and demonstrate its functionality on AdVectors with inhibitory genes. Aim 2 will test the hypothesis that a single siRNA can be broadly applied to inhibit transgene expression. It is anticipated that both strategies will be used in the ultimate cell line constructed in phase II. The final GMP manufacturing cell line will speed and facilitate the testing of advector vaccine concepts for many antigens, speed their progress to clinical testing and reduce the cost of goods for future approved vaccine products. [unreadable] [unreadable]